Ammonia, anhydrous

Endpoint summary

Key value for chemical safety assessment

Effects on fertility

Description of key information

No evidence of any reproductive effects was seen in reproductive screening and 2-generation reproductive toxicity studies with ammonium salts diammonium phosphate and ammonium perchlorate, respectively. The physiological role of ammonia indicates that it is unlikely to be a reproductive toxin at relevant exposure levels.

No additional studies are available.

Link to relevant study records

Reference

Reference 1

Endpoint:

screening for reproductive / developmental toxicity

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

See attached read-across justification

Reason / purpose for cross-reference:

read-across source

Limit test:

no

Clinical signs:

no effects observed

Body weight and weight changes:

effects observed, treatment-related

Food consumption and compound intake (if feeding study):

effects observed, treatment-related

Organ weight findings including organ / body weight ratios:

no effects observed

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Other effects:

effects observed, treatment-related

Reproductive function: oestrous cycle:

not examined

Reproductive function: sperm measures:

not examined

Reproductive performance:

no effects observed

Treatment at all dosages was well tolerated and there were no reatment-related deaths. A dosage dependent increase in transient post-dosing
salivation was apparent, which was considered to be due to the palatability of the test formulations rather than toxicity. A dosage-dependent increase in the number of animals with reddening of the extremities was also apparent mainly during the early stages of treatment.

Body weight gain and food consumption of males at 1500 mg/kg bw/day appeared to be suppressed when compared with the control group, such that gain between weeks 0-5 for this group was 78% of controls. The body weight gain for reproductive subgroup females receiving 1500 mg/kg bw/day was reduced during the first week of gestation, after which the values returned to levels comparable with the control.

Some treatment-related effects on hematology were evident (reduction in activated partial thromboplastin time for males at 750 and 1500 mg/kg bw/day, a non dosage-dependent elevation of alkaline phosphatase levels at 750 and 1500 mg/kg bw/day, reduced glucose and phosphorous levels at 1500 mg/kg bw/day, a dosage-dependent reduction in total protein at 750 and 1500 mg/kg bw/day with a slight elevated albumin/globulin ratio at the top dosage. Changes in females were limited to a decrease in phosphorous levels and a non-significant increase in alkaline phosphatase level at 1500 mg/kg bw). Relative kidney and liver weights for females at 1500 mg/kg bw/day were greater than in the control group, but there were no histological changes associated.

A number of treated animals at 750 and 1500 mg/kg bw/day exhibited horizontal banding on the incisors at necropsy; histological processing of these tissues failed to detect any change in the areas examined suggesting that the banding was restricted to the enamel of the teeth. The only histological findings related to treatment were the inflammatory/degenerative stomach changes in all treated groups that were considered likely to have arisen due to an irritant effect of the test formulations. There were no changes apparent at behavioural testing.

Mating performance and fertility were unaffected by treatment. There were no effects on the time to achieve conception, and pregnancy length.


Summary of effects on reproduction/development (control, low, mid, high dose):
Females achieving pregnancy: n= 9, 10, 10, 10.
Dams with live young born: n = 9, 10, 10, 10.
Implants/dam (mean): 15.7, 15.7, 14.1, 15.4.
Live pups/dam at birth (mean): 14.8, 14.6, 12.7, 14.0.
Live pups/dam at day 4 (mean): 14.6, 14.3, 12.7, 14.0.
sex ratio (% m) at birth (mean): 54.2, 52.7, 50.0, 48.5.
sex ratio (%m) at day 4 (mean): 54.2, 53.0, 50.0, 48.5.
Male pup weight at birth (mean): 6.4, 6.3, 6.6, 6.3.
Male pup weight at day 4 (mean): 8.7, 8.5, 9.2, 8.7.
Female pup weight at birth (mean): 5.9, 6.0, 6.1, 6.0.
Female pup weight at day 4 (mean): 8.2, 7.9, 8.6, 8.4.
Post-Implantation survival index: 95.2, 93.5, 90.0, 94.8.
Live birth index: 99.3, 99.4, 100.0, 95.9.
Viability index: 98.6, 98.2, 100.0, 100.0.

Dose descriptor:

NOAEL

Effect level:

1 500 mg/kg bw/day (nominal)

Based on:

other: Diammonium phosphate

Sex:

male/female

Basis for effect level:

other: Reproduction/developmental toxicity

Dose descriptor:

LOAEL

Effect level:

> 1 500 mg/kg bw/day (nominal)

Based on:

other: Diammonium phosphate

Sex:

male/female

Basis for effect level:

other: Reproduction/developmental toxicity

Clinical signs:

no effects observed

Mortality / viability:

no mortality observed

Body weight and weight changes:

no effects observed

Sexual maturation:

not examined

Organ weight findings including organ / body weight ratios:

not examined

Gross pathological findings:

no effects observed

Histopathological findings:

not examined

Parental treatment had no apparent effect on the offspring to day 4 of age; necropsy findings were unremarkable

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

1 500 mg/kg bw/day

Based on:

other: diamomonium phosphate

Sex:

male/female

Basis for effect level:

other: reproductive & developmental toxicity

Reproductive effects observed:

not specified

There were no treatment-related deaths and no signs of overt clinical toxicity. Bodyweight gain for reproductive subgroup females receiving 1,500 mg/kg/day was reduced during the first week of gestation (82% of control), after which they returned to levels comparable to the controls for the remainder of the study. Mating performance and fertility were unaffected by treatment, and parental treatment had no apparent effect on the offspring to day 4 of age.

Conclusions:

Diammonium phosphate NOAEL: 1,500 mg/kg/day (reproduction/developmental toxicity)
Diammonium phosphate LOAEL: >1,500 mg/kg/day (reproduction/developmental toxicity)

Executive summary:

The toxicity of read-across substance diammonium phosphate (DAP) was assessed in a combined repeated dose toxicity study with a reproduction / developmental toxicity screening test in rats. The NOAEL was found to be: 250 mg/kg/day (general toxicity); 1,500 mg/kg/day (reproduction/developmental toxicity). The LOAEL was found to be: 750 mg/kg/day (general toxicity); >1,500 mg/kg/day (reproduction/developmental toxicity). Mating performance and fertility were unaffected by treatment, and parental treatment had no apparent effect on the offspring up to age 4 days.

Reference 2

Endpoint:

two-generation reproductive toxicity

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

weight of evidence

Justification for type of information:

See attached read-across justification

Reason / purpose for cross-reference:

read-across source

Limit test:

no

Clinical signs:

no effects observed

Body weight and weight changes:

no effects observed

Food consumption and compound intake (if feeding study):

no effects observed

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Histopathological findings: non-neoplastic:

effects observed, treatment-related

Other effects:

effects observed, treatment-related

Reproductive function: oestrous cycle:

no effects observed

Reproductive function: sperm measures:

no effects observed

Reproductive performance:

no effects observed

P Generation Adults
The actual consumed doses met or exceeded target doses, doses consumed during gestation exceeded target doses by 33 -34% in all groups. This was attributed to the higher water consumption values of dams during pregnancy.
There were no deaths, abortions, or premature deliveries attributed to exposure. No clinical observations were considered related to exposure because there were no dose-response relationships, and were observations that were noted commonly in the laboratory with this strain of rat.
Absolute (g/day) and relative (g/kg/day) water consumption values for male rats in both 0.3 and 30.0 mg/kg.day dose groups were significantly reduced (p ≤ 0.05), compared to controls. In contrast, average maternal absolute and relative water consumption values during the precohabitation, gestation and lactation periods were comparable among the four groups. Food consumption (absolute and relative) for male and female rats was generally unaffected by treatment. Average body weights for male or female rats (during the precohabitation, gestation, and lactation periods) were comparable among the four exposure groups throughout the study.
Absolute thyroid weights were increased for male rats in all three treatment groups; the increases were significant at 3 and 30 mg/kg. Absolute thyroid weights were significantly increased in the 30 mg/kg females. The ratios of thyroid weight to terminal body weight and to the absolute brain weight were significantly increased in both males and females of the 30 mg/kg group. Absolute and relative weights for the other organs collected at necropsy were comparable to control values.
Mating and fertility paramaters in P generation males were unaffected by exposure. There were no statistically significant effects on sperm motility, sperm count, sperm density or morphology.
Oestrus cycling, mating and fertility parameters were unaffected in P generation females. Natural delivery observations were unaffected by exposure, the number of dams delivering litters, gestation duration, averages for implantations, live litter sizes and stillbirths, dams with stillborn pups, viability and lactation indices, sex ratios and pup body weights were comparable across all four groups.
Exposure-related histomorphologic changes were observed only in the thyroid gland. These changes occurred in males and females and were primarily hypertrophy and hyperplasia of the thyroid epithelium. All other microscopic changes observed in other organs were considered to have occurred spontaneously and be typical of those that occur in rodent reproductive studies.
TSH levels were significantly increased, and T4 levels signficantly decreased in 30 mg/kg males. Significant increases in serum T3 levels occurred in the 0.3 and 3.0 mg/kg males. TSH, T3 and T4 were unaffected by treatment in females.

F1 Generation Adults
The average actual consumed daily doses were 88 -100% of the target for males, and 67 -100% of the target for females, during all periods except lactation. During lactation, the F1 females were exposed to 116 -133% of target doses.
There were no deaths, abortions, or premature deliveries. All clinical observations that occurred in the F1 generation rats were considered unrelated to exposure. Absolute relative water and food consumption values were unaffected by exposure. No treatment related effects were observed at necropsy. Terminal body weights for the F1 males were significantly increased in the 0.3 mg/kg group, but this was not considered to be related to treatment as there was no dose relationship. Terminal body weights for females were unaffected by treatment.
Thyroid weights and the ratios of the thyroid weight to the terminal body weight and to the brain weight were significantly increased for males in the 3.0 and 30 mg/kg groups, and for females in the 0.3, 3.0 and 30 mg/kg groups.
Mating and fertility in F1 males were comparable between control and treated groups. The fertility index and the number of rats pregnant per rats in cohabitation were significantly increased in the 0.3, 3.0 and 30 mg/kg groups compared to the controls. This result was not thought to be related to treatment because there was no dose response relationship and was probably due to lower fertility in the control group. There were no significant effects on sprem motility, sperm count, morphology or density.
Oestrus cycling observations were comparable among treated females prior to cohabitation. Observations in natural delivery and lactation were unaffected by treatment. There was no effect on the number of days in cohabitation or the number of rats mating during the first or second week of cohabitation.
Exposure related histomorphologic changes were observed in the thyroid gland of the adult male and female F1 rats. These changes were primarily hypertrophy and hyperplasia of the thyroid follicular epithelium. The incidence was significantly increased in males from the 3 and 30 mg/kg groups and females from the 30 mg/kg group.

Dose descriptor:

NOAEL

Effect level:

30 mg/kg bw/day (nominal)

Sex:

male/female

Basis for effect level:

other: Excluding perchlorate-specific effects on the thyroid

Clinical signs:

no effects observed

Mortality / viability:

no mortality observed

Body weight and weight changes:

no effects observed

Sexual maturation:

no effects observed

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Gross pathological findings:

no effects observed

Histopathological findings:

effects observed, treatment-related

F1 Generation Pups
All clinical and necropsy observations on the F1 generation pups were considered unrelated to exposure.
The weight of the thyroid was significantly increased for both male and female pups in the 30 mg/kg group. The weight of the spleen was slightly increased in all treated groups, but the increase was only significant in the 30 mg/kg females.
Exposure-related histomorphologic changes were observed in the thyroid gland of male and female F1 pups. The changes were primarily hypertrophy and hyperplasia of the thyroid follicular epithelium. The incidence was significantly increased in males and females at 30 mg/kg, and in females at 3 mg/kg.
Serum T3 was significantly reduced in 30 mg/kg females. TSH levels were significantly reduced for males in the 0.3 and 3.0 mg/kg groups. Serum T4 levels were significantly increased for females in the 0.3 mg/kg.
There were no effects on the average day of preputial separation in F1 males. The average day of vaginal patency for treated F1 females was comparable to controls. The percentages of liveborn pups and still born pups were significantly decreased and increased, respectively, in the 30 mg/kg group. These changes were not thought to be treatment related because the average number of liveborn and stillborn pups per litter were comparable in the four exposure groups.

F2 Generation Pups
All clinical and necropsy observations in the F2 pups were considered unrelated to treatment. Thyroid weight was significantly increased for the females in the 30 mg/kg group. There were no other effects on organ weights. Exposure related histomorphologic changes were observed in the thyroid gland of male and female F2 pups, the incidence of hyperplasia/hypertrophy was significantly increased in the 3 and 30 mg/kg groups. Thyroid and pituitary hormone levels were unaffected by parental exposure.

Dose descriptor:

NOAEL

Generation:

F1

Effect level:

30 mg/kg bw/day (nominal)

Based on:

other: ammonium perchlorate

Sex:

male/female

Basis for effect level:

other: Excluding perchlorate-specific effects on the thyroid

Dose descriptor:

NOAEL

Generation:

F2

Effect level:

30 mg/kg bw/day (nominal)

Based on:

other: ammonium perchlorate

Sex:

male/female

Basis for effect level:

other: Excluding perchlorate-specific effects on the thyroid

Reproductive effects observed:

not specified

There were no effects on reproduction in any generation, therefore the NOAEL can be considered to be 30 mg/kg/day.

Specific exposure periods are not detailed in the paper.

Conclusions:

Ammonium perchlorate administration in the drinking water did not have an adverse effect on reproduction in the rat.

Executive summary:

This two-generation reproductive study examines the effects of read-across substance ammonium perchlorate on the male and female reproductive systems in rats, and on the growth and development of offspring. Adult Sprague-Dawley rats (30/sex/group) were given continuous access to ammonium perchlorate in their drinking water at doses of 0, 0.3, 3.0, and 30.0 mg/kg/day. F1 generation rats were given the same ammonium perchlorate doses as their respective P1 generation sires and dams beginning at weaning and continuing through the day of sacrifice. Standard reproductive parameters were evaluated; blood was collected for determination of serum thyroid-stimulating hormone (TSH), triiodothyronine (T3), and thyroxine (T4) levels. Histopathological examination was conducted on major tissues, including the thyroid. No significant changes in developmental parameters were observed. In the F1 generation adult rats, relative thyroid weights were significantly increased in all dose groups for female rats and in the 3.0 and 30.0 mg/kg/day dose groups for male rats. Histopathologic changes in the thyroid consisted of hypertrophy and hyperplasia that increased in incidence and severity in a dose-related manner. Dose-related, statistically significant changes in TSH and T4 or T3 occurred at doses higher than those that resulted in changes in thyroid weight and thyroid histopathology, 30 mg/kg/day. Thus, perchlorate is not a reproductive toxicant in rats when administered in the drinking water at doses up to 30 mg/kg/day, but it can affect the thyroid at doses of 3 mg/kg/day and above. Based on the perchlorate-specific effects on the thyroid, the authors suggested a 0.3 mg/kg-day for the no-observable-adverse-effect level (NOAEL).

Effect on fertility: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

387 mg/kg bw/day

Study duration:

subacute

Species:

rat

Quality of whole database:

Two studies are available on read-across substances: one screening study using diammonium phosphate and one two generation study using ammonium perchlorate

Effect on fertility: via inhalation route

Endpoint conclusion:

no study available

Effect on fertility: via dermal route

Endpoint conclusion:

no study available

Additional information

Screening study

The potential for the read-across substance diammonium phosphate to cause reproductive toxicity in rats has been investigated in a four week reproductive/developmental toxicity screening test conducted according to OECD Test Guideline 422 (HLS, 2002). In the study groups of rats were exposed to the substance at 0, 250, 750 and 1500 mg/kg bw/day via oral gavage. No evidence of any effects on fertility or foetal development were seen at dose levels up to 1500 mg/kg bw/day; this dose level is equivalent to ‘target’ NOAEL of 0.258 x 1500 mg/kg = 387 mg/kg bw/day ammonia.

 

Two-generation study

The potential for ammonium perchlorate to cause reproductive toxicity in rats has been investigated in a two generation study conducted according to OECD Test Guideline 416 (York et al, 2001). In the study, rats were exposed to the substance at concentrations of 0, 0.3, 3.0 and 30 mg/kg bw/day via their drinking water. No effects on reproductive parameters were observed at dose levels of up to and including 30 mg/kg bw/day. While the study did identify effects on the parental thyroid associated with perchlorate exposure, these findings were not attributable to ammonium. The results of the study therefore confirm that exposure to ammonium is not associated with reproductive toxicity.

 

Conclusion

There is no evidence that exposure to ammonium ions will cause reproductive toxicity. Inhalation exposure to ammonia will result in an equilibrium in the blood (at physiologically relevant pH) between non-ionised ammonia (NH3) and ionised ammonium (NH4+) in a ratio of approximately 1:100. The same equilibrium will exist in animals orally exposed to ammonium salts, therefore read-across is appropriate. Human maternal blood contains measurable levels of ammonia as a consequence of protein catabolism; the blood in the hepatic portal circulation contains much higher levels of ammonia due to its generation from urea by the gastrointestinal flora. Ammonia is rapidly and effectively detoxified in the liver by the urea cycle and also via additional pathways, therefore will not accumulate and is unlikely to cause any reproductive effects.

Effects on developmental toxicity

Description of key information

No evidence of any developmental effects have been seen in rabbits and rats in studies with ammonia or ammonium salts ammonium chloride, ammonium perchlorate and ammonium acetate. The physiological role of ammonia indicates that it is unlikely to be a developmental toxin at relevant exposure levels.

Link to relevant study records

Reference

Reference 1

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Non-guideline farm animal reproduction study

Qualifier:

no guideline followed

Principles of method if other than guideline:

Inhalation exposure in pigs with some examinations of reproductive and developmental toxicity

GLP compliance:

not specified

Remarks:

: non-standard, published study

Limit test:

no

Species:

pig

Strain:

other: crossbred: Yorkshire x Hampshire x Chester White

Details on test animals or test system and environmental conditions:

Crossbred gilts (young female pigs) were reared in a conventional grower unit from 2 to 4.5 months of age, where they were naturally exposed to mycoplasmal and bacterial pathogens that cause pneumonia and atrophic rhinitis. At 4.5 months of age, gilts were moved to environmentally regulated rooms and randomly assigned to one of two treatment groups.

Route of administration:

inhalation: vapour

Type of inhalation exposure (if applicable):

whole body

Vehicle:

unchanged (no vehicle)

Details on exposure:

"Aerial ammonia" at 7±1 ppm (low) and 35±3 ppm (moderate) dose levels.
The low dose was obtained by flushing manure pits weekly. The moderate dose was obtained by adding anhydrous ammonia to manure pits that were not flushed.

Analytical verification of doses or concentrations:

yes

Details on analytical verification of doses or concentrations:

Air samples were obtained weekly in each room, using a volumetric pump. Ammonia concentration was determined by use of direct-read colour-detector tubes. Replicate air samples were obtained at 0.3 m above the floor at random locations in each room, and mean concentration was determined. When ammonia was added to the manure pit, aerial ammonia concentration was also obtained after ammonia discharge.

Details on mating procedure:

After 6 weeks exposure half the gilts were exposed daily to mature boars and bred at first oestrus.

Duration of treatment / exposure:

Pigs were exposed for 6 weeks then half were slaughtered and half were bred. Bred pigs continued to be exposed until day 30 of gestation.

Frequency of treatment:

Daily

Duration of test:

Pigs were exposed for 6 weeks then half were slaughtered and half were bred. Bred pigs continued to be exposed until day 30 of gestation.

Remarks:

Doses / Concentrations:
7±1 and 35±3 ppm
Basis:
analytical conc.

No. of animals per sex per dose:

40 gilts (young females) per dose: 20 from each dose were bred.

Control animals:

yes, concurrent no treatment

Details on study design:

Prior to ammonia exposure (whilst housed in a conventional grower), the pigs were naturally exposed to mycoplasmal and bacterial pathogens that cause pneumonia and atrophic rhinitis. After 6 weeks of exposure, half of each group were slaughtered and the other half were bred. The bred gilts continued to be exposed until day 30 of gestation. A limited number of developmental parameters were then tested.

Maternal examinations:

Gilts were weighed biweekly. Respiratory tissue was examined after slaughter. Age of puberty and conception rate were recorded.

Ovaries and uterine content:

The number of live foetuses, foetal weight and foetal length were recorded at day 30 of gestation.

Fetal examinations:

The number of live foetuses, foetal weight and foetal length were recorded at day 30 of gestation.

Statistics:

Least squares ANOVA, using general linear model procedures.

Indices:

Not determined.

Historical control data:

No information available

Details on maternal toxic effects:

Maternal toxic effects:yes

Details on maternal toxic effects:
Gilts of the moderate exposure group showed a lower mean daily weight gain in the first two weeks of exposure (n=40). After six weeks of exposure (n=40) and at day 30 of gestation (n=20), a reduction in body weight was found in this group (significant after 6 weeks of exposure (n=20). At both exposure levels, animals slaughtered after 6 weeks of exposure showed lung lesions and moderate degeneration of nasal turbinates. Weight at puberty was lower for moderate dose group gilts compared to the low dose (109.7 kg cf. 118.2 kg, respectively).

Dose descriptor:

NOEL

Effect level:

> 35 ppm

Basis for effect level:

other: developmental toxicity

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
At day 30 of gestation, there was no difference in the number of live foetuses or in foetal weight and length between the two dose groups.

Dose descriptor:

NOAEC

Effect level:

35 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

other: no effects in fetuses

Abnormalities:

not specified

Developmental effects observed:

not specified

No significant differences in ovarian weight and uterine weight were observed between the low and moderate dose groups slaughtered after 6 weeks of exposure. There was no differences between the groups of age at puberty (day of breeding) and conception rate.

Conclusions:

Aerial ammonia exposure up to ~35 ppm for 6 weeks prior to breeding and continuing until day 30 of gestation did not appear to be toxic to the developing foetus. Maternal toxicity as evidenced by a reduction in weight gain was apparent at 35 ppm, but not 7 ppm.

Executive summary:

Gilts were exposed to to 7 ppm or 35 ppm atmospheric ammonia for 6 weeks prior to breeding and continuing until day 30 of gestation. Ammonia exposure does not appear to be toxic to the developing foetus. Maternal toxicity as evidenced by a reduction in weight gain was apparent at 35 ppm, but not 7 ppm. At both exposure levels, animals slaughtered after 6 weeks of exposure showed lung lesions and moderate degeneration of nasal turbinates. However, the value of this study is limited because of the limited number of development parameters tested and the absence of control animals.

Reference 2

Endpoint:

developmental toxicity

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

See attached read-across justification

Reason / purpose for cross-reference:

read-across source

Species:

rabbit

Strain:

New Zealand White

Details on maternal toxic effects:

Maternal toxic effects:yes

Details on maternal toxic effects:
There were no treatment-related deaths. Two does in the 1 mg/kg group aborted on GD 28 and were sacrificed. Both abortions were considered unrealted to treatment because the incidences were not dose-dependent. One dam in the 100 mg/kg group prematurely delivered on GD 27. Because rabbits normally deliver on GD 31, and the pups appeared to be full term (they had fur and were nursing), it was assumed that the rabbit had been incorrectly identified and shipped by the supplier on the wrong day of gestation.
Clinical observations included localised alopecia, ungroomed coat, scant soft or liquid faeces, and a red perivaginal, perinasal or perioral substance. The clinical observations recorded were considered unrelated to treatment because the incidences were not dose-dependent; the observation was associated with abortion of a litter, and/or the observations were commonly seen in rabbits in the laboratory environment.
The maternal body weights in the control group were consistently lower than the treated groups over the gestation period. There were however, no statistically significant differences in average maternal body weights, gravid uterine weights, body weight gains, or corrected GD29 body weights (GD29 bodyweight minus the gravid uterine weight) among exposure groups during gestation.
The only adverse necropsy observation was a mottled liver that occurred in the 1 mg/kg group doe that aborted.
Microscopic examination of the dams' thyroid glands revealed hypertrophy of the follicular epithelium in the 10, 30 and 100 mg/kg groups, this was considered to be treatment related by the authors. No treatment related microscopic changes were observed in the thyroid gland of any rabbits from the 0.1 or 1.0 mg/kg groups. In the affected thyroids, there was an increased height or enlargement of the follicular epithelium, occasionally resulting in a decrease in the lumen of follicles, which contained pale and occasionally vacuolated colloid.
Serum T4 levels appeared to decrease in a treatment-related manner, with the 30 and 100 mg/kg groups showing statistically significant decreases compared to controls. This decrease was considered to be treatment-related by the authors because it corresponded to the hypertrophy of follicular epithelium. There were no statistically signficant changes in serum T3 and TSH in treated rabbits compared to controls.

Dose descriptor:

NOAEL

Effect level:

100 mg/kg bw/day (nominal)

Based on:

other: ammonium perchlorate

Basis for effect level:

other: developmental toxicity

Dose descriptor:

NOAEL

Effect level:

1 mg/kg bw/day (nominal)

Based on:

other: ammonium perchlorate

Basis for effect level:

other: maternal toxicity

Details on embryotoxic / teratogenic effects:

Embryotoxic / teratogenic effects:no effects

Details on embryotoxic / teratogenic effects:
Caesarean section observations were based on 22, 24, 23, 24 and 23 pregnant does surviving to GD 29. There were no effects on caesarean sectioning or litter parameters. All values were within laboratory historical ranges. All placentae appeared normal and no dam had a litter consisting of only resorbed conceptuses. Litter averages for corpora lutea, implantations, litter sizes, live and dead foetuses, percent dead or resorbed conceptuses, and foetal body weights were comparable and did not differ significantly between groups.
Foetal evaluations were based on 180, 184, 196, 195, 189 and 206 live foetuses in the six exposure groups, respectively. No foetal alterations were attributable to treatment. Only 6 foetuses had gross external alterations: 3 from the control group and two from the 1 mg/kg group, and 1 from the 100 mg/kg group litters. Foetal soft tissue alterations only occurred in four foetuses of the control group, two in the 0.1 mg/kg group, three in the 1 mg/kg groups, two in the 30 mg/kg group and five in the 100 mg/kg group.
A statistically significant difference in incidence of folded retina was observed; the foetal incidence in all the treated groups was lower than the control group. Folded retina of the right and/or left eye occurred in several foetuses, but was only seen in the heads examined with Wilson;s technqieu and was therefore considered to be an artefact of processing. Foetuses in some of the exposure groups showed significant increases in skeletal alterations, however none of the changes were considered to be treatment-related as there was no evidence of a dose-response relationship.

Dose descriptor:

NOAEL

Effect level:

100 mg/kg bw/day

Based on:

other: ammonium perchlorate

Sex:

male/female

Basis for effect level:

other: No effects

Abnormalities:

not specified

Developmental effects observed:

not specified

Actual consumed doses of ammonium perchlorate on GDs 6 to 19 were 0, 0.1, 1.0, 10.8, 33.9 and 114.2 mg/kg/day. During GDs 19 to 29, actual consumed doses of ammonium perchlorate were 0, 0.1, 0.8, 10.0, 26.7 and 90.4 mg/kg/day. Therefore, average actual consumed doses for the entire period of gestation were 0, 0.1, 0.9, 10.4, 30.3 and 102.3 mg/kg/day.

Conclusions:

There was no evidence that ammonium perchlorate is toxic to the developing foetus. An increased incidence of thyroid follicular hypertrophy was observed in does, however the thyroid is a target organ for the perchlorate ion therefore this effect is not attributed to the ammonium ion.

Executive summary:

This developmental toxicity study was conducted to evaluate the embryo-foetal toxicity and teratogenic potential of read-across substance ammonium perchlorate in New Zealand White [Hra:(NZW)SPF] rabbits. Pregnant rabbits were given continual access to ammonium perchlorate in drinking water at target doses of 0, 0.1, 1.0, 10.0, 30.0, and 100.0 mg/kg/day on gestation days 6 through 28. The actual consumed doses in the study were 0, 0.1, 0.9, 10.4, 30.3, and 102.3 mg/kg/day. The rabbits were sacrificed on gestation day 29, and foetuses were examined for developmental alterations. In addition, blood was collected from does for determination of serum thyroid stimulating hormone (TSH), triiodothyronine (T3), and thyroxine (T4) levels and the thyroid was subjected to histopathologic examination. No maternal deaths were attributed to perchlorate exposure. Ammonium perchlorate as high as 100.0 mg/kg-day did not affect caesarean sectioning or litter parameters studied, and all values were found to be within the historical ranges of the laboratory. The litter averages for corpora lutea, implantations, litter sizes, live and dead foetuses, percent dead or resorbed conceptuses, and fetal body weights were comparable and also did not differ significantly in the six dose groups. All placentae appeared normal and no dam had a litter consisting of only resorbed conceptuses. The maternal thyroid was the target organ for ammonium perchlorate in this study. Increased incidence of thyroid follicular hypertrophy was observed in does treated with 10 mg/kg/day and above, and significantly decreased T4 was observed in does treated with 30 mg/kg/day and above. Based on these data, the maternal no-observable-adverse-effect level (NOAEL) for ammonium perchlorate was 1.0 mg/kg-day. The developmental NOAEL for ammonium perchlorate was found to be 100.0 mg/kg-day for rabbits.

Effect on developmental toxicity: via oral route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEL

14.5 mg/kg bw/day

Study duration:

subacute

Species:

rabbit

Quality of whole database:

Three published studies are available

Effect on developmental toxicity: via inhalation route

Endpoint conclusion:

no adverse effect observed

Dose descriptor:

NOAEC

25 mg/m³

Study duration:

subchronic

Species:

pig

Quality of whole database:

Only one study available

Effect on developmental toxicity: via dermal route

Endpoint conclusion:

no study available

Additional information

No evidence of developmental toxicity was seen in a key rabbit guideline-compliant pre-natal developmental toxicity study (OECD Test Guideline 414) conducted using ammonium perchlorate at doses up to highest dose level of 100 mg/kg bw/day (York et al, 2001). Maternal toxicity was demonstrated by increased incidence of thyroid follicular hypertrophy in does treated dosed with 10 mg/kg/day and above, and significantly decreased T4 was observed in does treated with 30 mg/kg/day and above. Based on these data, the maternal NOAEL for ammonium perchlorate was 1.0 mg/kg-day. The source developmental NOAEL of 100 mg/kg bw (ammonium perchlorate) is equivalent to ‘target’ NOAEL of 0.145 x 100 mg/kg = 14.5 mg/kg bw/day ammonia.

 

In a supporting non-guideline-compliant pre-natal developmental toxicity study, rats were exposed to drinking water containing 0 and 0.17 mol/L ammonium chloride (i.e., 3060 mg NH4 +/L) from gestation day 7 (Goldman & Jakovac, 1964) . Signs of embryotoxicity noted were decreased foetal weights and crown-rump lengths. No foetal skeletal or gross pathology abnormalities were found. Maternal mortality was the only parameter measured for dams, so it is unclear whether the exposure level was high enough to affect the foetus and drinking water consumption was not reported.

 

The influence of ammonium ions on foetal development was investigated in mice in a supporting non-standard study involving in vitro exposure prior to transplantation into dams (Lane & Gardner, 1994). Examination on gestational day 15 showed an apparent relationship between the duration of exposure and the incidence of exencephaly. Embryos that were cultured with various concentrations of ammonium ion before being transferred to recipient dams showed increased incidence of exencephaly and a decreased percentage of implantation sites with increased ammonium concentration. It is unclear how embryos might be exposed to ammonia or ammonium in vivo or if in vivo exposure would affect foetal development and implantation in a way similar to that described in this study.

 

Finally, no evidence of foetal toxicity was seen in a supporting study in pigs exposed to maternally toxic concentrations of 7 and 35 ppm ammonia by inhalation for 6 weeks prior to breeding and continuing until day 30 of gestation (Diekman et al, 1993). Although the design of the study is somewhat limited, it can be concluded that the relatively low concentrations of ammonia required to induce local irritant effects evoked maternal toxicity as evidenced by a reduction in weight gain at 35 ppm but no developmental toxicity. The concentration of 35 ppm, equivalent to 25 mg/m³ can be considered as a developmental NOAEC in pigs.

 

Conclusion

There is no evidence that exposure to ammonium ions causes specific developmental toxicity in vivo. Inhalation exposure to ammonia will result in an equilibrium in the blood (at physiologically relevant pH) between non-ionised ammonia (NH3) and ionised ammonium (NH4+) in a ratio of approximately 1:100. The same equilibrium will exist in animals orally exposed to ammonium salts, therefore read-across is appropriate. Human maternal blood contains measurable levels of ammonia as a consequence of protein catabolism; levels of ammonia in foetal blood are slightly higher. The blood in the hepatic portal circulation contains much higher levels of ammonia due to its generation from urea by the gastrointestinal flora. Ammonia is rapidly and effectively detoxified in the liver by the urea cycle and also via additional pathways, therefore will not accumulate and is unlikely to cause any developmental toxic effects at relevant exposure levels.

Justification for classification or non-classification

The available data do not indicate that ammonia is a reproductive or developmental toxin. Classification of the substance for reproductive or developmental toxicity according to CLP Regulation 1272/2008/EC is therefore not warranted.